Sensitivity: The minimum detectable dose of this kit is typically less than 27.4pg/mL.

Specifity: This assay has high sensitivity and excellent specificity for detection of High Temperature Requirement Factor A4 (HTRA4).
No significant cross-reactivity or interference between High Temperature Requirement Factor A4 (HTRA4) and analogues was observed.

Stability: The stability of kit is determined by the loss rate of activity. The loss rate of this kit is less than 5% within the expiration date under appropriate storage condition.
To minimize extra influence on the performance, operation procedures and lab conditions, especially room temperature, air humidity, incubator temperature should be strictly controlled. It is also strongly suggested that the whole assay is performed by the same operator from the beginning to the end.

Storage: Store at 4°C for frequent use. Stored at -20°C in a manual defrost freezer for one year without detectable loss of activity. Avoid repeated freeze-thaw cycles.

Notes: For In vitro laboratory use only. Not for any clinical, therapeutic, or diagnostic use in humans or animals. Not for animal or human consumption.

Recovery

Matrices listed below were spiked with certain level of recombinant High Temperature Requirement Factor A4 (HTRA4) and the recovery rates were calculated by comparing the measured value to the expected amount of High Temperature Requirement Factor A4 (HTRA4) in samples.

Matrix

Recovery range (%)

Average(%)

serum(n=5)

78-96

86

EDTA plasma(n=5)

77-95

82

heparin plasma(n=5)

77-97

90

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level High Temperature Requirement Factor A4 (HTRA4) were tested 24 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level High Temperature Requirement Factor A4 (HTRA4) were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

Linearity

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of High Temperature Requirement Factor A4 (HTRA4) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Test principle

The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to High Temperature Requirement Factor A4 (HTRA4). Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to High Temperature Requirement Factor A4 (HTRA4). Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain High Temperature Requirement Factor A4 (HTRA4), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of High Temperature Requirement Factor A4 (HTRA4) in the samples is then determined by comparing the O.D. of the samples to the standard curve.